Heat shock proteins are involved in a wide variety of processes, both physiological and pathological. Heat shock protein 27 (HSP27) is a member of the small heat shock protein family, which comprises members ranging from 15 to 30 kDa in size and which may be phosphorylated or oligomerized under various conditions. HSP27 is principally described as an intracellular chaperone capable of binding and stabilizing the actin cytoskeleton in response to stress. In addition, HSP27 can bind cytochrome c and prevent downstream caspase activation, making it a potent anti-apoptotic protein. More recently, it was discovered that HSP27 can interact with estrogen receptor β and reduce transcriptional signaling through the estrogen response element1. Although multi-faceted, the functions described for HSP27 have been solely thought to be within the confines of the cell membrane; however, extracellular release of HSP27 may be regulated and may have important effects on steps leading to the development of atherosclerosis.
While the role of estrogens in atherosclerotic coronary artery disease has enjoyed intense scrutiny over the past two decades or more, recent reappraisals of clinical trials and new experimental data strongly argue for a second look at the role of not only “hormone replacement therapy” (HRT), but also novel hormone manipulation strategies for the prevention/attenuation of atherosclerosis. There are several important caveats to clinical studies of HRT that help explain the absence of the expected cardiovascular benefits of HRT in post-menopausal women2-4. A principal deficiency of these studies is the late introduction of HRT to women, who, for example, in a Women's Health study, were on average 7 years post-menopausal before HRT was commenced5, 6. Many of these women may have been susceptible to the potential ravages of atherosclerosis while deficient of ovarian hormones in the initial years post-menopause. Hence, when HRT was introduced, it may have been too late to derive an “atheroprotective” (or reversal of atherosclerosis) effect.
The biological effects of estrogen are mediated by at least two cellular receptors: ER alpha (ERα) and ER beta (ERβ) that belong to the classical steroid hormone receptor superfamily. When activated, the receptors translocate to the nucleus and modulate transcriptional activity through interactions with estrogen response elements (EREs). These receptors also participate in signaling cascades at the cell membrane, suggestive of a function entirely independent of gene regulation7. Structurally, ERα and ERβ are subdivided into several functional domains including ligand binding, DNA binding, and both ligand-independent (AF-1) and ligand-dependent (AF-2) activation domains. While the two receptors share considerable structural similarities, they derive functional specificity via differential tissue expression patterns and regions of structural diversity (e.g. the A/B domain where there is only 30% sequence identity between ERβ and ERα)8, 9. Moreover, ER ligand complexes produce different effects in different cells due to variable expression of co-regulatory proteins (e.g. co-activators and co-repressors). Indeed, in some instances the physiological and pathophysiological response to hormones may reside with these co-regulatory molecules, rather than solely with the receptors themselves.
Approximately 300 nuclear receptor (NR) associated proteins are known, typically as a result of yeast two hybrid screens that employ the NR as “bait” (cf. review by Smith and O'Malley)10. In general, these proteins do not bind DNA directly, but instead facilitate the interaction of hormone receptors with DNA and other structural proteins—ultimately serving to facilitate (activator) or hinder (repressor) the activation of transcription.
For a variety of reasons ERβ has emerged as a key receptor in the vessel wall. For example, the expression of ERβ mRNA is markedly up-regulated after vascular injury in male arteries11, 12. Moreover, in male arteries, ERβ is the predominant receptor expressed in the intima, media and adventitia, and its expression correlates with the degree of calcification—a marker of severe atherosclerosis. Therefore, ERβ appears to play an important yet unidentified role in the progression of atherosclerosis. Yeast two-hybrid analysis revealed that HSP27 is an ERβ associated protein. HSP27 attenuated ERβ transcriptional activity preserved endothelial cell homeostasis, and normal volunteers had >3-fold higher serum levels than those patients with angiographic evidence of coronary artery disease.
It has been observed that HSP27 may be a potential biomarker for atherosclerosis, with expression of HSP27 diminishing with the progression of disease1, 13, 14. Serum levels of HSP27 have been shown to be attenuated in patients with atherosclerosis compared to healthy individuals13. HSP27 may be involved in long term vessel wall homeostasis that is then lost with the progression of atherosclerosis. Although the mechanisms by which HSP27 may be “atheroprotective” are not yet elucidated, many believe that analogous to its effects in other tissues (e.g. nerve, gastromucosal and myocardium) it protects the vessel wall from stressful stimuli and prevents apoptosis15, 16. While this may in part explain why HSP27 levels have been shown to be acutely increased in the serum following myocardial ischemia14, HSP27 also appears to be involved in the long-term maintenance of vessel wall homeostasis that unfortunately may be lost with the progression of CAD.
It is, therefore, desirable to provide a method for preventing or treating cardiovascular disease using HSP27.